Our long-term objective is to determine how extra-cellular signals are sensed by the cell cycle machine and then transmitted into regulated cell cycle progression. The specific aims of this proposal are to define the role o the site-specific phosphorylation in the regulation of cyclin D1 nuclear import and export, to determine the relationship between cyclin D1 localization and the proteolysis, to explore the potential oncogenicity of cyclin D1 proteins that are constitutively nuclear, and to determine the mechanism of cyclin D1-dependent transformation. Interspecies heterokaryon assays will be used to examine the parameters that regulate cyclin D1 nuclear export, and in vitro nuclear import assays will be used to directly determine the mechanism(s) of cyclin D1 nuclear import. The necessity of nuclear export for efficient cyclin Dl proteolysis will also be determined using both in vitro and in vivo techniques. Finally, we will test the hypothesis that failure to remove cyclin Dl from the nucleus during S-phase is critical for normal cellular proliferation by creating mice carrying the cyclin D1 variant, D 1-T286A, whose expression is controlled by the immunoglobulin intron enhancer element, E(mu). The subversion of normal growth signaling pathways is a hallmark of neoplasia, and the capacity of cyclin D1 to act as a mitogenic sensor that integrates growth factor signals into cell cycle progression makes it a frequent target in Cancer. The studies proposed will identify the regulatory mechanisms that determine cyclin D1 subcellular localization, determine the role of localization in cyclin D1 accumulation, and demonstrate how cells constrain cyclin D1 activity, thereby preventing neoplasia.